Scientists at the National Center for Natural Products Research at the University of Mississippi have developed a system they say improves upon existing methods for testing mosquito repellents in a laboratory setting. Shown above, the system includes a 30- by 30- by 30-cm aluminum cage with an acrylic reservoir containing human blood, warmed to human body temperature by a water circulator below it. Over the blood, a sheet of collagen mimics human skin. (Photo credit: Abbas Ali, Ph.D.)

By Edward Ricciuti

Among the processes by which researchers gauge the effectiveness of a mosquito repellent is one that depends on convincing a volunteer to shove an arm treated with the compound into a box and let the little bloodsuckers have a go at it. The system works well enough but, like all such assays, has limitations, not the least of which is finding volunteers. Due to odors and other variable skin characteristics, moreover, not all humans provoke the same response by mosquitoes, creating troublesome variables.

Ed Ricciuti

Another method uses a heat source to mimic a warm body and collagen membrane as a substitute for human skin, placed over a container holding human blood, within what amounts to a small plastic box. It is effective, but the small size of the module inhibits mosquito flight, an indicator of repellence, as well as the number of mosquitoes used during a test.

Given the global threat of malaria, dengue fever, Zika and other diseases transmitted by mosquitoes, the quest for a new wave of repellents and ways to evaluate them goes on apace. Scientists at the National Center for Natural Products Research at the University of Mississippi have developed an assay system that they say offers many benefits over the in vivo test using human subjects and is more effective than other in vitro, or artificial, test methods as well. Their new system is described in research published in the Journal of Medical Entomology (JME).

“Finding new repellents that are safe for human use and the environment is a big challenge for researchers,” say the project scientists. “Many researchers around the globe are screening materials to find new compounds for protecting human beings from mosquito bites. Evaluations of mosquito repellents both in the laboratory and the field can help establish the best repellent materials, application rates, and residual effects of the active materials.”

Not all repellents are equal. Some evaporate quickly from skin or clothing. Others can be diluted by sweat or water. Some work only in massive doses. Repellents such as DEET (diethyltoluamide), which has been on the market for more than 50 years, can have side effects.

“Evaluations of mosquito repellents both in the laboratory and the field can help establish the best repellent materials, application rates, and residual effects of the active materials,” the researchers write in JME.

The complicated processes involved in the development of the new assay method illustrate a point sometimes forgotten by the public at large: Research and development of ways to test new biomedical products can be as exhaustive as those needed to create it. The assay test engineered by the University of Mississippi scientists builds upon methods already in use.

“To minimize the involvement of human subjects and to get meaningful data, we have developed and standardized a new large cage A & K bioassay system that is efficient, accurate, and has the capability to do an increased number of replications in a shorter period of time with limited resource,” write the authors of the paper, Abbas Ali, Ph.D., Charles L. Cantrell, Ph.D., and Ikhlas A. Khan, Ph.D. The method is known as the “A & K” bioassay system, after two of the authors.

The A & K system, tested on Aedes aegypti mosquitoes, consists of a 30-by-30-by-30-centimeter collapsible aluminum cage with metal screens and a transparent acrylic viewing panel. It is spacy enough to hold a large number of mosquitoes at one time, reducing the number of tests required. Importantly, it has room enough for mosquitoes to easily fly, a critical behavior when assaying repellents.

When conducting trial runs of the equipment, researchers released a couple of hundred mosquitoes into the cage and watched them react to human blood in an acrylic reservoir, warmed to human body temperature by a water circulator below it. Over the blood, a sheet of collagen mimicked human skin.

After one minute, they disturbed the mosquitoes. Those that were not feeding already were or went airborne when roused. Feeders remained drawing blood through the collagen. If only one percent or less—two or fewer—of the mosquitoes continued to feed, the repellent was considered to be at its maximum effective dose. The researchers noted that when the test was done without repellent, as a control, mosquitoes quickly landed on the collagen. When repellents themselves were tested, the time it took for mosquitoes to land—or whether they landed at all—depended on the potency of the compound. Based on the results of tests with the A & K system, researchers can assess the dosage of repellents to be used on humans.

“We are confident that this bioassay will prove to be an excellent tool to be used extensively by researchers who are working on the screening and testing of compounds for repellency against mosquitoes,” the authors of the paper conclude.

Ed Ricciuti is a journalist, author, and naturalist who has been writing for more than a half century. His latest book is called Bears in the Backyard: Big Animals, Sprawling Suburbs, and the New Urban Jungle (Countryman Press, June 2014). His assignments have taken him around the world. He specializes in nature, science, conservation issues, and law enforcement. A former curator at the New York Zoological Society, and now at the Wildlife Conservation Society, he may be the only man ever bitten by a coatimundi on Manhattan’s 57th Street.